STRUCTURE/FUNCTION OF PITSLRE KINASES

PITSLRE蛋白激酶是一种CDK，包括20多个 不同的异构体，并普遍表达。人们对他们的情况知之甚少 功能。PITSLRE p110激酶亚型定位于核质和 核斑点，由染色质内颗粒团(IGCs)组成的区域， 它们是转录/RNA加工因子的存储和组装部位。 申请人已经发现PITSLRE p110激酶直接与RNPS1相互作用， 通用RNA剪接因子和转录延伸因子EKK2， 在体外和体内。PITSLRE p110还与RNAPII和CKII在 活着。CKII使不完全重复序列的特定部分磷酸化 RNAPII CTD的区域以及氨基末端的特定区域 PITSLRE p110的体外结构域。目前的提案将解决 假设PITSLRE p110激酶调控转录、RNA加工或 和与CKII的关联在这些功能和/或 这种激酶在细胞核内的定位。PITSLRE p110复合体将 进行生化提纯，并从核质和免疫球蛋白中鉴定。 它的底物和调节器将被确定。申请者将决定 如果CKII在体内磷酸化PITSLRE p110和CTD，并评估 磷酸化的生物学后果。最后，各种角色的作用 PITSLRE激酶亚型将通过敲除小鼠的基因来检测。

The PITSLRE protein kinases are CDKs that include more than 20 different isoforms and are ubiquitously expressed. Little is known about their function. The PITSLRE p110 kinase isoforms are localized to the nucleoplasm and nuclear speckles, regions composed of intrachromatin granule clusters (IGCs), which are storage and assembly sites for transcription/RNA processing factors. The applicant has found that PITSLRE p110 kinase interacts directly with RNPS1, a general RNA splicing factor, and EKK2, a transcriptional elongation factor, in vitro and in vivo. PITSLRE p110 also associates with RNAPII and CKII in vivo. CKII phosphorylates a very specific portion of the imperfect repeat region of the RNAPII CTD as well as a specific region of the amino terminal domain of PITSLRE p110 in vitro. The current proposal will address the hypothesis that PITSLRE p110 kinases regulate transcription, RNA processing, or both and that its association with CKII plays a role in these functions an/or the localization of this kinase within the nucleus. PITSLRE p110 complexes will be biochemically purified and characterized from the nucleoplasm and the IGC. Its substrates and regulators will be identified. The applicant will determine if CKII phosphorylates PITSLRE p110 and the CTD in vivo and also assess the biological consequences of the phosphorylation. Finally the role of the various PITSLRE kinase isoforms will be examined by knocking-out the genes in mice.